Myocytes from adult normal and diabetic rats will be isolated by a procedure which we have developed in our laboratory. Metabolic intermediates, associated with glucose utilization and energy production will be determined to correlate the metabolic status of the different cellular preparations. Changes in glucose uptake and glucose metabolism in the normal and diabetic cells, in response to insulin will be correlated with alteration of insulin binding to cells from diabetic animals. Binding experiments will be designed to determine changes in receptor population, affinity constants and possible compartmentalization of the hormone. Possible effects of insulin on calcium transport will be investigated, by determining changes in total calcium levels and alterations in 45Ca2 ion fluxes of normal and diabetic cells. The stimulation of Na/K-ATPase system by insulin, will be evaluated, in order to assess any possible interactions between this process and calcium uptake. Finally, the molecular regulation by insulin and the effects of diabetes on calmodulin and/or calcium will be investigated in the intact heart and heart cells. Calmodulin will be purified from rat testes and used to determine its presence in the normal and diseased states. Identification of the protein will be made by gel filtration and gel electrophoresis in polyacrylamide gels. In addition, the effects of insulin administration "in vivo" on calmodulin levels during diabetes will be determined as well as the possible inhibitory effect of cyclohexamide. Similar experiments will be undertaken "in vitro" to determine possible regulation of calmodulin by calcium and insulin in the isolated myocytes. The insulin-dependent phosphorylation of several proteins will be assayed "in vitro" in normal and diabetic heart cells, and compared to results obtained from calmodulin stimulated phosphorylation. The possibility that the intermediate effects of insulin may be regulated by calmodulin-calcium-insulin interactions will be investigated.